problems with negative control

Hi,
I did co-ip with mouse ab and mouse IgG as negative control.blot it and prob with different ab but again mouse.
I'm getting very nice band at the expected size compare to the control, but i can also see it, less strong, in the igG
is this still can be positive and how can I reduce it
Thanks
Michal

Have you done the Preclearing with the Beads (Protein G i guess for Mouse) ?? this would reduce the background in Negtive control..Try out this and see if you havent done. and increase washing steps, this might also help...

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Hi,I did co-ip with mouse ab and mouse IgG as negative control.blot it and prob with different ab but again mouse.I'm getting very nice band at the expected size compare to the control, but i can also see it, less strong, in the igG is this still can be positive and how can I reduce itThanksMichal

I would prefer being perfectionist rather than a passionist in Research.

I did the precleaning with the beads and increased the washes without change,I'm using the protein a/g agarose beads from santa cruzI heard that there is secondary ab that do not recognize the Igg, does anyone know what does it mean?ThanksM

Have you done the Preclearing with the Beads (Protein G i guess for Mouse) ?? this would reduce the background in Negtive control..Try out this and see if you havent done. and increase washing steps, this might also help...

.

Hi,I did co-ip with mouse ab and mouse IgG as negative control.blot it and prob with different ab but again mouse.I'm getting very nice band at the expected size compare to the control, but i can also see it, less strong, in the igG is this still can be positive and how can I reduce itThanksMichal